Pyridazin-3(2H)-one derivatives as monoamine oxidase selective isoform B inhibitors

ABSTRACT

The present invention refers to pyridazin-3(2H)-one derivatives of general structure I, II and III, which are selective MAO-B inhibitors, and to the use thereof for preparing medicaments intended to treat disorders derived from MAO-B hyperactivity, particularly degenerative disorders of the central nervous system (CNS), such as Parkinson&#39;s disease (PD), Alzheimer&#39;s disease (AD) and other dementias. These are pyridazin-3(2H)-one derivatives having dithiocarbamate moieties bonded to position 4, 5 or 6 through an alkyl chain of variable length (n=1, 2, 3). This invention is also directed to the preparation of said compounds.

CROSS REFERENCE TO RELATED APPLICATION

This Application is a 371 of PCT/ES2015/000029 filed on Mar. 3, 2015,which claims the benefit of Spanish Patent Application No. P201400162filed on March 2014, both of which are incorporated herein by reference.

TECHNICAL FIELD

The present invention refers to novel C4-, C5- or C6-substitutedpyridazinone derivative compounds having a dithiocarbamate moiety, andof general structure I, II and III, respectively, which are selectiveMAO-B inhibitors, and to the use thereof for preparing medicamentsintended to treat disorders derived from MAO-B hyperactivity,particularly degenerative disorders of the central nervous system (CNS),such as Parkinson's disease (PD), Alzheimer's disease (AD) and otherdementias.

These are pyridazin-3(2H)-one having dithiocarbamate moieties bonded toposition 4, 5 or 6 through an alkyl chain of variable length. Thegeneral structural formulae of the 3 series of compounds, structures I,II and III, are detailed below.

State of the Art

The monoamine oxidases (MAO) are flavoenzymes being present in the outermembrane of CNS cells and peripheral tissue mitochondria, wherein theycatalyse the oxidative deamination of endogenous or exogenous amines soas to generate the corresponding aldehydes, ammonia and H₂O₂. Two MAOisoenzymes are known, designated as MAO-A and MAO-B, which shareapproximately 70% of the amino acid sequence and which aredifferentiated by the three dimensional structure thereof, by thesubstrate selectivity and by the existence of selective inhibitors(Proc. Natl. Acad. Sci. USA 105, 5739-5744, 2008; J. Biol. Chem.280(16), 15761-15766, 2005). Both isoenzymes play an important role inthe regulation of biogenic amines concentration in the brain; this fact,together with the substrate selectivity, determines the clinicalimportance of the MAO inhibitors (MAOIs). Thus, the MAO-A shows higheraffinity for serotonin (5-hydroxytryptamine, 5-HT), adrenaline (A) andnoradrenaline (NA), and it is selectively inhibited by clorgiline andmoclobemide, whereas MAO-B preferably degrades 3-phenylethylamine andbenzylamine and it is selectively inhibited by selegiline andrasagiline. There are some MAOIs which feature lack of selectivity, suchas iproniazid.

The structure of the MAOI compounds cited above is detailed in figure 1.

Functional studies about both enzymes have revealed that MAOs play animportant role in the regulation of biogenic amines concentration in thebrain, which are involved in different pathological processes affectingthe CNS, which determines the clinical importance of MAOIs (Curr. Med,Chem. 11, 2033-2043, 2004). MAO-A inhibition in the CNS enhancesnoradrenaline and serotonin levels, two neurotransmitters involved indepressive disorders, whereas MAO-B inhibition increase dopamine levels,which in PD are reduced, which explains that MAOI-A are used asantidepressants and anxiolytics, and MOIs-B for EP treatment.

The AD is a progressive neurodegenerative disease which is the mosthabitual type of senile dementia. Although the aetiology thereof ismultiple and complex, it is associated to β-amyloid plaques (βA) in thebrain, which can promote the loss of cholinergic neurons in the cerebralcortex and in the hippocampus, which explains the cognitive deficiencyand memory loss manifesting in the short term in patients undergoing AD(Velázquez Farmacología Básica y Clínica 17 ed. Panamericana: Madrid2005, 329-335). Therefore, traditional pharmacological treatment of ADinvolves administration of acetylcholinesterase inhibitors (Rang y DaleFarmacologia 6 ^(a) ed. Elsevier: Barcelona 2008, 515-516), an enzymewhich degrades acetylcholine. However, studies have been done whichevidence an increasing activity of MAO-B in the brain of patientsundergoing certain neurodegenerative disorders such as, for example, PDor AD (Biochem. Pharmacol. 38, 555-561, 1989) and new therapeuticexpectations have arisen. MAO-B activity increase originates an increasein the reactive oxygen species (ROS) which contribute to oxidativestress and neuron death. Although more studies are required forclarifying the beneficial effects of MAOI-B in neurodegenerativeprocesses such as AD, said effects are related to ROS reduction, whichis neurotoxic, and with monoamines increase in the brain of thesepatients (Neurotoxicology 25, 271-277, 2004; Journal of NeuroscienceResearch 79, 172-179, 2005).

Currently, the main therapeutic application of MAOI-B is in PD treatment(Translational Neurodegeneration 1:10, 2012; TranslationalNeurodegeneration 2:19, 2013), a neurological disorder which affectsmotor activity and results from a decrease in striatum dopamine levels,caused by progressive death of nigrostriatal neurons. Although theclassical treatment of the PD have been administration of L-dopa(precursor of dopamine) associated to an inhibitor of peripheral dopadecarboxylase enzyme, more recent therapeutic alternatives involveadministration of catechol ortho-methyl transferase inhibitors (COMT),such as entacapone, and also MAOI-B selective inhibitors, such asselegiline and rasagiline (Translational Neurodegeneration 1:10, 2012).

There are several articles and patents describing compounds which act asselective inhibitors of MAO-B and applications thereof inneurodegenerative disorders, such as for example derivatives of coumarin(ES 2343347; J. Med. Chem. 54, 7127-7131, 2011) (compound 1, figure 2),γ-chromones (Bioorg. Med. Chem. Lett. 20, 2709-2712, 2010; Bioorg. Med.Chem. Lett. 21, 707-709, 2011) (compound 2, figure 2), pyrazolines andother diazaheterocycle derivatives (J. Med. Chem. 48, 7113-7122, 2005;Bioorg. Med Chem. Lett 20, 6479-6482, 2010; J. Med. Chem. 49, 3743-3747,2006; J. Med Chem. 50, 5364-5371, 2007) (compounds 3 and 4, figure 2),thiazolyl-hydrazines (J. Med. Chem. 53, 6516-6520, 2010; Arch. Pharm.Chem. Life Sci. 346, 17-22, 2013) (compound 5, figure 2),dithiolane-thiones (WO2006/089861) (compound 6, figure 2), and amines oramides derived from heterocyclic systems (EP 1524267; WO 2004/007429;EP1524265, J. Med. Chem. 50, 922-931, 2007) (compounds 7 and 8, figure2).

Figure 2 shows a detailed structure of several compounds having MAOI-Bactivity.

Pyridazine is a diazine which is rare in natural products. However, thisheteronucleus is part of a small group of structures known asprivileged, due to the capacity thereof of generating compounds beingactive against several targets, (Med. Chem. Comun. 2, 935-941, 2011).Pyridazine derivatives have a wide spectrum of pharmacological activity(cardiotonic, anti-hypertensive, platelet antiaggregate, hypolipidemic,analgesic and anti-inflammatory, antinociceptive, anti-depressant,anxiolytic, GABA antagonist, hypoglycaemic, anti-infectious orantineoplastic, among others), and many of them are analogues to thestructure of 3(2H)-pyridazinone (Progress in Medicinal Chemistry,Elsevier Science Publishers Biomedical Division: Amsterdam 1990, 1-49;Progress in Medicinal Chemistry, Elsevier Science Publishers BiomedicalDivision: Amsterdam 1992, 141-183; Med. Chem. Res 22, 2539-2552, 2013).

The pyridazine ring is present in compounds acting as MAO-B selectiveinhibitors; these are condensed polycyclic systems (J. Med. Chem. 49,3743-3747, 2006; J. Med Chem. 50, 5364-5371, 2007; J. Med. Chem. 49,6264-6272, 2007) (compound 4, figure 2). Furthermore, there are articlesand patents referring to simple pyridazine derivatives which act uponother therapeutic targets being efficient in neurodegenerativedisorders, such as, for example, agonists of GABA_(A) receptor (WO2012/068161; WO 2010/127968) (compound 9, figure 3), agonists of thecannabinoid receptor CB2 (WO 2011/097553) (compound 10, figure 3),activators of glutamate transporter protein (WO 2013/019938) (compound11, figure 3), modulators of γ-secretase (Med. Chem. Lett. 1, 184-187,2010; Bioorg. Med Chem. Lett. 21, 4016-4019, 2011) (compound 12, figure3), or inhibitors of tau protein oligomerization (Biochemistry, 48,7732-7745, 2009) (compound 13, figure 3), some of which are3-(2H)-pyridazinone (compounds 10, 12 and 13).

However, 3-(2H)-pyridazinone derivatives which act as selective MAOI-Bare not known.

The compounds of the present invention lack of structural relationshipwith those described so far, and behaved as selective inhibitors againstMAO-B. These are novel 3-(2H)-pyridazinone derivatives substituted inpositions 4, 5 or 6 with dithiocarbamate moieties, bonded to saidpositions through an alkyl chain of variable length, which selectivelyinhibit MAO-B activity when the bioactivity thereof is assayed in vitro.

DESCRIPTION OF THE INVENTION

The present invention refers to novel C4-, C5- or C6-substitutedpyridazinone derivative compounds having dithiocarbamate moieties, andof general structure I, II and III, respectively, which are selectiveMAO-B inhibitors in vitro, and to the possible use thereof for preparingmedicaments intended to treat disorders derived from MAO-Bhyperactivity, particularly degenerative disorders of the centralnervous system (CNS), such as Parkinson's disease (PD), Alzheimer'sdisease (AD) and other dementias.

These are pyridazin-3(2H)-one having dithiocarbamate moieties bonded toposition 6 through an alkyl chain of variable length and of the generalformula I.

wherein,

-   -   n is an integer number selected from 1, 2, 3, 4, 5, 6, 7, 8;    -   R is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a C₁-C₆ carboxyalkyl group, a C₁-C₆ haloalkyl group, a        C₆-C₁₂ aryl group, a C₆-C₁₂ aralkyl group, a C₄-C₁₂ heteroaryl        group;    -   R¹ is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a halogen atom,    -   R² is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a halogen atom,    -   R³, R⁴, being the same or different, are selected from: a        hydrogen atom, a C₁-C₆ alkyl group, saturated C₁-C₆        heterocycloalkyl group, a C₆-C₁₂ aryl group, a C₆-C₁₂ aralkyl        group, a C₄-C₁₂ heteroaryl group,    -   Or R³ and R⁴ form a cycle selected from: C₅-C₈ cycloalkyl, C₅-C₈        heterocycloalkyl, N-alkyl substituted C₅-C₈ heterocycloalkyl,        N-aryl substituted C₅-C₈ heterocycloalkyl, N-cycloalkyl        substituted C₅-C₈ heterocycloalkyl, N-aralkyl substituted C₅-C₈        heterocycloalkyl, N-acyl substituted C₅-C₈ heterocycloalkyl.

And preferably

-   -   R is CH₃, phenyl (Ph) or benzyl (Bn).    -   R¹ is H, halogen (Cl, Br, I) or an alkyl chain    -   R² is hydrogen (H) or methyl (CH₃).    -   n is optionally 1, 2 or 3.    -   R³ and R⁴ may be hydrogen, alkyl groups being the same or        different, such as methyl (CH₃) or ethyl (CH₂CH₃), or, together        with the nitrogen atom (N), they may constitute a 5 or 6        membered heterocyclic ring, being aliphatic or incorporating an        oxygen atom (O) or a second N atom. This second N atom may be        substituted with a linear (CH₃, CH₂CH₃) or cyclic (cyclopropyl)        alkyl group, or with an aryl group (Ph), aralkyl (Bn) or aroyl        (benzoyl, Bz).

In a particular aspect, the compounds of the general formula I arerepresented by the formulae Ia₁-a₃₃ (table I), Ib₁-b₃₃ (table II),Ic₁-c₃₃ (table III) and Id₁-d₃₃ (table IV), wherein R¹ is preferably H.

TABLE I Ia

n = 1 n = 2 n = 3 —N(CH₃)₂ Ia₁ Ia₁₂ Ia₂₃ —N(CH₂CH₃)₂ Ia₂ Ia₁₃ Ia₂₄

Ia₃ Ia₁₄ Ia₂₅

Ia₄ Ia₁₅ Ia₂₆

Ia₅ Ia₁₆ Ia₂₇

Ia₆ Ia₁₇ Ia₂₈

Ia₇ Ia₁₈ Ia₂₉

Ia₈ Ia₁₉ Ia₃₀

Ia₉ Ia₂₀ Ia₃₁

Ia₁₀ Ia₂₁ Ia₃₂

Ia₁₁ Ia₂₂ Ia₃₃

TABLE II Ib

n = 1 n = 2 n = 3 —N(CH₃)₂ Ib₁ Ib₁₂ Ib₂₃ —N(CH₂CH₃)₂ Ib₂ Ib₁₃ Ib₂₄

Ib₃ Ib₁₄ Ib₂₅

Ib₄ Ib₁₅ Ib₂₆

Ib₅ Ib₁₆ Ib₂₇

Ib₆ Ib₁₇ Ib₂₈

Ib₇ Ib₁₈ Ib₂₉

Ib₈ Ib₁₉ Ib₃₀

Ib₉ Ib₂₀ Ib₃₁

Ib₁₀ Ib₂₁ Ib₃₂

Ib₁₁ Ib₂₂ Ib₃₃

TABLE III Ic

n = 1 n = 2 n = 3 —N(CH₃)₂ Ic₁ Ic₁₂ Ic₂₃ —N(CH₂CH₃)₂ Ic₂ Ic₁₃ Ic₂₄

Ic₃ Ic₁₄ Ic₂₅

Ic₄ Ic₁₅ Ic₂₆

Ic₅ Ic₁₆ Ic₂₇

Ic₆ Ic₁₇ Ic₂₈

Ic₇ Ic₁₈ Ic₂₉

Ic₈ Ic₁₉ Ic₃₀

Ic₉ Ic₂₀ Ic₃₁

Ic₁₀ Ic₂₁ Ic₃₂

Ic₁₁ Ic₂₂ Ic₃₃

TABLE IV Id

R = Me R = Ph R = Bn —N(CH₃)₂ Id₁ Id₁₂ Id₂₃ —N(CH₂CH₃)₂ Id₂ Id₁₃ Id₂₄

Id₃ Id₁₄ Id₂₅

Id₄ Id₁₅ Id₂₆

Id₅ Id₁₆ Id₂₇

Id₆ Id₁₇ Id₂₈

Id₇ Id₁₈ Id₂₉

Id₈ Id₁₉ Id₃₀

Id₉ Id₂₀ Id₃₁

Id₁₀ Id₂₁ Id₃₂

Id₁₁ Id₂₂ Id₃₃

These are pyridazin-3(2H)-one having dithiocarbamate moieties bonded toposition 5 through an alkyl chain of variable length and of generalformula II.

wherein,

-   -   n is an integer number selected from 1, 2, 3, 4, 5, 6, 7, 8;    -   R is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a C₁-C₆ carboxyalkyl group, a C₁-C₆ haloalkyl group, a        C₆-C₁₂ aryl group, a C₆-C₁₂ aralkyl group, a C₄-C₁₂ heteroaryl        group;    -   R¹ is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a halogen atom,    -   R² is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a halogen atom,    -   R³, R⁴, being the same or different, are selected from: a        hydrogen atom, a C₁-C₆ alkyl group, saturated C₁-C₆        heterocycloalkyl group, a C₆-C₁₂ aryl group, a C₆-C₁₂ aralkyl        group, a C₄-C₁₂ heteroaryl group,    -   Or R³ and R⁴ form a cycle selected from: C₅-C₈ cycloalkyl, C₅-C₈        heterocycloalkyl, N-alkyl substituted C₅-C₈ heterocycloalkyl,        N-aryl substituted C₅-C₈ heterocycloalkyl, N-cycloalkyl        substituted C₅-C₈ heterocycloalkyl, N-aralkyl substituted C₅-C₈        heterocycloalkyl, N-acyl substituted C₅-C₈ heterocycloalkyl.

And preferably

-   -   R is CH₃, phenyl (Ph) or benzyl (Bn).    -   R¹ is H, halogen (Cl, Br, I) or an alkyl chain    -   R² is hydrogen (H).    -   n is optionally 1, 2 or 3.    -   R³ and R⁴ may be hydrogen, alkyl groups being the same or        different, such as methyl (CH₃) or ethyl (CH₂CH₃), or, together        with the nitrogen atom (N), they may constitute a 5 or 6        membered heterocyclic ring, being aliphatic or incorporating an        oxygen atom (O) or a second N atom. This second N atom may be        substituted with a linear (CH₃, CH₂CH₃) or cyclic (cyclopropyl)        alkyl group, or with an aryl group (Ph), aralkyl (Bn) or aroyl        (benzoyl, Bz).

In a particular aspect, the compounds of general formula II arerepresented by formulae IIa₁-a₃₃ (table V), IIb₁-b₃₃ (table VI),IIc₁-c₃₃ (table VII) where R¹ and R² are preferably H.

TABLE V IIa

n = 1 n = 2 n = 3 —N(CH₃)₂ IIa₁ IIa₁₂ IIa₂₃ —N(CH₂CH₃)₂ IIa₂ IIa₁₃ IIa₂₄

IIa₃ IIa₁₄ IIa₂₅

IIa₄ IIa₁₅ IIa₂₆

IIa₅ IIa₁₆ IIa₂₇

IIa₆ IIa₁₇ IIa₂₈

IIa₇ IIa₁₈ IIa₂₉

IIa₈ IIa₁₉ IIa₃₀

IIa₉ IIa₂₀ IIa₃₁

IIa₁₀ IIa₂₁ IIa₃₂

IIa₁₁ IIa₂₂ IIa₃₃

TABLE VI IIb

n = 1 n = 2 n = 3 —N(CH₃)₂ IIb₁ IIb₁₂ IIb₂₃ —N(CH₂CH₃)₂ IIb₂ IIb₁₃ IIb₂₄

IIb₃ IIb₁₄ IIb₂₅

IIb₄ IIb₁₅ IIb₂₆

IIb₅ IIb₁₆ IIb₂₇

IIb₆ IIb₁₇ IIb₂₈

IIb₇ IIb₁₈ IIb₂₉

IIb₈ IIb₁₉ IIb₃₀

IIb₉ IIb₂₀ IIb₃₁

IIb₁₀ IIb₂₁ IIb₃₂

IIb₁₁ IIb₂₂ IIb₃₃

TABLE VII IIc

n = 1 n = 2 n = 3 —N(CH₃)₂ IIc₁ IIc₁₂ IIc₂₃ —N(CH₂CH₃)₂ IIc₂ IIc₁₃ IIc₂₄

IIc₃ IIc₁₄ IIc₂₅

IIc₄ IIc₁₅ IIc₂₆

IIc₅ IIc₁₆ IIc₂₇

IIc₆ IIc₁₇ IIc₂₈

IIc₇ IIc₁₈ IIc₂₉

IIc₈ IIc₁₉ IIc₃₀

IIc₉ IIc₂₀ IIc₃₁

IIc₁₀ IIc₂₁ IIc₃₂

IIc₁₁ IIc₂₂ IIc₃₃

These are pyridazin-3(2H)-one representing dithiocarbamate moietiesbonded to position 4 through an alkyl chain of variable length and ofthe general formula III.

wherein,

-   -   n is an integer number selected from 1, 2, 3, 4, 5, 6, 7, 8;    -   R is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a C₁-C₆ carboxyalkyl group, a C₁-C₆ haloalkyl group, a        C₆-C₁₂ aryl group, a C₆-C₁₂ aralkyl group, a C₄-C₁₂ heteroaryl        group;    -   R¹ is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a halogen atom,    -   R² is a group selected from: a hydrogen atom, a C₁-C₆ alkyl        group, a halogen atom,    -   R³, R⁴, being the same or different, are selected from: a        hydrogen atom, a C₁-C₆ alkyl group, saturated C₁-C₆        heterocycloalkyl group, a C₆-C₁₂ aryl group, a C₆-C₁₂ aralkyl        group, a C₄-C₁₂ heteroaryl group,    -   Or R³ and R⁴ form a cycle selected from: C₅-C₈ cycloalkyl, C₅-C₈        heterocycloalkyl, N-alkyl substituted C₅-C₈ heterocycloalkyl,        N-aryl substituted C₅-C₈ heterocycloalkyl, N-cycloalkyl        substituted C₅-C₈ heterocycloalkyl, N-aralkyl substituted C₅-C₈        heterocycloalkyl, N-acyl substituted C₅-C₈ heterocycloalkyl.

And preferably

-   -   R is CH₃, phenyl (Ph) or benzyl (Bn).    -   R¹ is H, halogen (Cl, Br, I) or an alkyl chain    -   R² is hydrogen (H).    -   n is optionally 1, 2 or 3.    -   R³ and R⁴ may be hydrogen, alkyl groups being the same or        different, such as methyl (CH₃) or ethyl (CH₂CH₃), or, together        with the nitrogen atom (N), they may constitute a 5 or 6        membered heterocyclic ring, being aliphatic or incorporating an        oxygen atom (O) or a second N atom. This second N atom may be        substituted with a linear (CH₃, CH₂CH₃) or cyclic (cyclopropyl)        alkyl group, or with an aryl group (Ph), aralkyl (Bn) or aroyl        (benzoyl, Bz).

In a particular aspect, the compounds of the general formula I arerepresented by formulae IIIa₁-a₃₃ (table VIII), IIIb₁-b₃₃ (table IX),IIIc₁-c₃₃ (table X), where R¹ and R² are preferably H.

TABLE VIII IIIa

n = 1 n = 2 n = 3 —N(CH₃)₂ IIIa₁ IIIa₁₂ IIIa₂₃ —N(CH₂CH₃)₂ IIIa₂ IIIa₁₃IIIa₂₄

IIIa₃ IIIa₁₄ IIIa₂₅

IIIa₄ IIIa₁₅ IIIa₂₆

IIIa₅ IIIa₁₆ IIIa₂₇

IIIa₆ IIIa₁₇ IIIa₂₈

IIIa₇ IIIa₁₈ IIIa₂₉

IIIa₈ IIIa₁₉ IIIa₃₀

IIIa₉ IIIa₂₀ IIIa₃₁

IIIa₁₀ IIIa₂₁ IIIa₃₂

IIIa₁₁ IIIa₂₂ IIIa₃₃

TABLE IX IIIb

n = 1 n = 2 n = 3 —N(CH₃)₂ IIIb₁ IIIb₁₂ IIIb₂₃ —N(CH₂CH₃)₂ IIIb₂ IIIb₁₃IIIb₂₄

IIIb₃ IIIb₁₄ IIIb₂₅

IIIb₄ IIIb₁₅ IIIb₂₆

IIIb₅ IIIb₁₆ IIIb₂₇

IIIb₆ IIIb₁₇ IIIb₂₈

IIIb₇ IIIb₁₈ IIIb₂₉

IIIb₈ IIIb₁₉ IIIb₃₀

IIIb₉ IIIb₂₀ IIIb₃₁

IIIb₁₀ IIIb₂₁ IIIb₃₂

IIIb₁₁ IIIb₂₂ IIIb₃₃

TABLE X IIIc

n = 1 n = 2 n = 3 —N(CH₃)₂ IIIc₁ IIIc₁₂ IIIc₂₃ —N(CH₂CH₃)₂ IIIc₂ IIIc₁₃IIIc₂₄

IIIc₃ IIIc₁₄ IIIc₂₅

IIIc₄ IIIc₁₅ IIIc₂₆

IIIc₅ IIIc₁₆ IIIc₂₇

IIIc₆ IIIc₁₇ IIIc₂₈

IIIc₇ IIIc₁₈ IIIc₂₉

IIIc₈ IIIc₁₉ IIIc₃₀

IIIc₉ IIIc₂₀ IIIc₃₁

IIIc₁₀ IIIc₂₁ IIIc₃₂

IIIc₁₁ IIIc₂₂ IIIc₃₃

In another aspect, the invention refers to a medicament comprising acompound of formula (I), (II) or (III), as it has been described above,or a salt thereof in a pharmaceutically acceptable carrier, having oneor more pharmaceutically acceptable excipients.

In a particular embodiment, said medicament also comprises one or moreadditional therapeutic agents.

Synthesis

The compounds Ia-d, IIa-c and IIIa-c could be obtained by means of anyknown chemical process being applicable to similar compounds.

In another aspect, the invention refers to a method for the synthesis ofa compound of formula (I), (II) or (III), as it has been describedabove, characterized in that it comprises at least a stage, wherein the6(5)(4)-bromoalkyl-3(2H)-pyridazinone of formula (IV), (IX) o (XIII), asecondary amine of formula V and carbon disulphide (CS₂) react in thepresence of a base in a solvent at room temperature.

The compounds Ia-d of the general formula I were obtained by means of amulti-component reaction between the 6-bromoalkyl-3(2H)-pyridazinones offormula IV, a secondary amine of formula V and carbon disulphide (CS₂)in the presence of anhydrous potassium phosphate (K₃PO₄) as an exampleof a base, in dimethylformamide (DMF) as an example of solvent, and atroom temperature (RT), such as it is shown in scheme 1.

Wherein R, R¹, R², R³, R⁴ and n are as described above for the compoundsof formula I.

CS₂ and amines of formula V are commercial compounds, while the6-bromoalkyl-3(2H)-pyridazinones of formula IV can be obtained from6-hydroxyalkyl-3(2H)-pyridazinones of formula VI, where R, R¹, R² and nare as described above, by carbon tetrabromide bromination (CBr₄) andtriphenylphosphine (PPh₃) or with N-bromosuccinimide (NBS) and PPh₃,adapting standard procedures (J. Heterocyclic Chem. 36, 985-990, 1999;Tetrahedron 50, 13575-13682, 1994).

Precursors of structure VI can be prepared in two stages (scheme 2) from5-(tert-butyldiphenylsyliloxyalkyl)-5-hydroxy(methoxy)-5H-furan-2-onesof structure VII, wherein R¹, R² and n are as described above, andsimilarly, as described in the bibliography (Bioorg. Med. Chem. Lett.20, 6624-6627, 2010; Magn. Reson. Chem., 49, 437-442, 2011). A firstreaction of the furanones VII with methylhydrazine (CH₃NHNH₂),phenylhydrazine (PhNHNH₂) or benzylhydrazine (BnNHNH₂) in ethanol(EtOH), provides the6-(tert-butyldiphenylsyliloxyalkyl)-3(2H)-pyridazinones of structureVIII, wherein R, R¹, R² and n are as described above, which transforminto the 6-hydroxyalkyl-3(2H)-pyridazinones VI by reaction withtetrabutylammonium fluoride (TBAF) in tetrahydrofuran (THF).

Furanones of structure VII can be prepared from the corresponding2-alkylfuranes by oxidation with singlet oxygen, in a manner beinganalogous to that described in the bibliography (Tetrahedron Lett. 45,5207-5209, 2004; Bioorg. Med. Chem. Lett. 20, 6624-6627, 2010; Magn.Reson. Chem., 49, 437-442, 2011).

Compounds IIa-c of general formula II were obtained by multicomponentreaction between 5-bromoalkyl-3(2H)-pyridazinones of formula IX, asecondary amine of formula V and carbon disulphide (CS₂) in the presenceof anhydrous potassium phosphate (K₃PO₄), in dimethylformamide (DMF) andat room temperature (RT), as it is shown in scheme 3.

Wherein R, R¹, R², R³, R⁴ and n are as described above for the compoundsof formula I.

CS₂ and amines of formula V are commercial compounds, whereas5-bromoalkyl-3(2H)-pyridazinones of formula IX can be obtained from5-hydroxyalkyl-3(2H)-pyridazinones of formula X, wherein R, R¹, R² and nare as described above, by carbon tetrabromide bromination (CBr₄) andtriphenylphosphine (PPh₃) or with N-bromosuccinimide (NBS) and PPh₃,adapting standard procedures (J. Heterocyclic Chem. 36, 985-990, 1999;Tetrahedron 50, 13575-13682, 1994).

Precursors of structure X can be prepared in two stages (scheme 4) from4-(tert-butyldiphenylsyliloxyalkyl)-5-hydroxy-5H-furan-2-ones ofstructure XI, wherein R¹, R² and n are as described above, andsimilarly, as described in the bibliography (Bioorg. Med. Chem. Lett.20, 6624-6627, 2010; Magn. Reson. Chem., 49, 437-442, 2011). A firstreaction of the furanones XI with methylhydrazine (CH₃NHNH₂),phenylhydrazine (PhNHNH₂) or benzylhydrazine (BnNHNH₂) in ethanol(EtOH), provides the5-(tert-butyldiphenylsyliloxyalkyl)-3(2H)-pyridazinones of structureXII, wherein R, R¹, R² and n are as described above, which transforminto the 5-hydroxyalkyl-3(2H)-pyridazinones X by reaction withtetrabutylammonium fluoride (TBAF) in tetrahydrofuran (THF).

Furanones of structure XI can be prepared from the corresponding3-alkylfuranes by oxidation with singlet oxygen, in a manner beinganalogous to that described in the bibliography (Tetrahedron Lett. 45,5207-5209, 2004; Bioorg. Med. Chem. Lett. 20, 6624-6627, 2010; Magn.Reson. Chem., 49, 437-442, 2011).

Compounds IIIa-c of general formula III were obtained by multicomponentreaction between 4-bromoalkyl-3(2H)-pyridazinones of formula XIII, asecondary amine of formula V and carbon disulphide (CS₂) in the presenceof anhydrous potassium phosphate (K₃PO₄), in dimethylformamide (DMF) andat room temperature (RT), as it is shown in scheme 5.

Where R, R¹, R², R³, R⁴ and n, are as described above for the compoundsof formula III.

CS₂ and amines of formula V are commercial compounds, whereas4-bromoalkyl-3(2H)-pyridazinones of formula XIII can be obtained from4-hydroxyalkyl-3(2H)-pyridazinones of formula XIV, wherein R, R¹, R² andn are as described above, by carbon tetrabromide bromination (CBr₄) andtriphenylphosphine (PPh₃) or with N-bromosuccinimide (NBS) and PPh₃,adapting standard procedures (J. Heterocyclic Chem. 36, 985-990, 1999;Tetrahedron 50, 13575-13682, 1994).

Precursors of structure XIV can be prepared in two stages (scheme 6)from 3-(tert-butyldiphenylsyliloxyalkyl)-5-hydroxy-5H-furan-2-ones ofstructure XV, wherein R¹, R² and n are as described above, andsimilarly, as described in the bibliography (Bioorg. Med. Chem. Lett.20, 6624-6627, 2010; Magn. Reson. Chem., 49, 437-442, 2011). A firstreaction of the furanones XV with methylhydrazine (CH₃NHNH₂),phenylhydrazine (PhNHNH₂) or benzylhydrazine (BnNHNH₂) in ethanol(EtOH), provides the5-(tert-butyldiphenylsyliloxyalkyl)-3(2H)-pyridazinones of structureXVI, wherein R, R¹, R² and n are as described above, which transforminto the 5-hydroxyalkyl-3(2H)-pyridazinones XIV by reaction withtetrabutylammonium fluoride (TBAF) in tetrahydrofuran (THF).

Furanones of structure XV can be prepared from the corresponding3-alkylfuranes by oxidation with singlet oxygen, in a manner beinganalogous to that described in the bibliography (Tetrahedron Lett. 45,5207-5209, 2004; Bioorg. Med. Chem. Lett. 20, 6624-6627, 2010; Magn.Reson. Chem., 49, 437-442, 2011).

Compounds of formula Ia-d, IIa-c and IIa-c selectively inhibit MAOisoform B and may be used for preparing medicaments intended to treatdisorders derived from MAO-B hyperactivity, as degenerative disorders ofthe central nervous system (CNS), such as Parkinson's disease (PD),Alzheimer's disease (AD) and other dementias.

Some representative compounds of formula Ia-d, IIa-c and IIIa-c to whichthe present invention refer to, are the following,

-   a) 1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethyl    N,N-diethyldithiocarbamate (Id₂).-   b) 1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethyl    Pyrrolidin-1-ylcarbodithioate (Id₃).-   c) 1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethyl    Piperidin-1-ylcarbodithioate (Id₄).-   d) 1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethyl    morpholin-4-ylcarbodithioate (Id₅).-   e) 1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethyl    4-benzoylpiperazin-1-ylcarbodithioate (Id₁₁).-   f) 2-(1-methyl-6-oxo-1,6-dihydropyridazin-3-yl)ethyl    Piperidin-1-ylcarbodithioate (Ia₁₅).-   g) 3-(1-methyl-6-oxo-1,6-dihydropyridazin-3-yl)propyl    Pyrrolidin-1-ylcarbodithioate (Ia₂₅).-   h) 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethyl    N,N-diethyldithiocarbamate (IIc₂).-   i) 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethyl    Pyrrolidin-1-ylcarbodithioate (IIc₃).-   j) 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethyl    Piperidin-1-ylcarbodithioate (IIc₄).-   k) 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethyl    Morpholin-4-ylcarbodithioate (IIc₅).-   l) 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethyl    4-benzoylpiperazin-1-ylcarbodithioate (IIc₁₁).-   m) 1-benzyl-6-oxo-1,6-dihydropyridazin-5-ylmethyl    Pyrrolidin-1-ylcarbodithioate (IIIc₃).-   n) 1-benzyl-6-oxo-1,6-dihydropyridazin-5-ylmethyl    Morpholin-4-ylcarbodithioate (IIIc₅).-   o) 1-benzyl-6-oxo-1,6-dihydropyridazin-5-ylmethyl    4-benzoylpiperazin-1-ylcarbodithioate (IIIc₁₁).

EXAMPLES

The examples given below should be considered as a way of providing abetter understanding of the present invention, without being limitativethereof.

General Procedures

Proton nuclear magnetic resonance spectra (¹H NMR) are in all casesaccording to the structures disclosed. The ¹H NMRs were registered inthe Bruker 400 DPX and Bruker ARX 400 spectrophotometer, usingdeuterated chloroform (CDCl₃) or deuterated methanol (CD₃OD). Chemicalshifts are expressed in δ units, in parts per million (ppm), relative totetramethylsilane (TMS), coupling constants (J) are indicated in Hertzs(Hz), and multiplicity as follows: s, singlet; d, doublet; t, triplet;m, multiplet. High resolution mass spectrometry (HRMS) was performed ina Bruker Microtof Focus spectrometer, using electrospray ionization(ESI) or electron impact ionization (EI).

Reactions under inert atmosphere were performed under argon (Ar)atmosphere. All the commercial reagents were directly taken from thebottles provided by the supplier and were used without being purified.Organic solvents were dried by means of standard procedures (Vogel'sTextbook of Practical Organic Chemistry 5th ed. Longman Scientific andTechnical: London 1989; Perrin, D. D., Armarego, W. L. F. Purificationof Laboratory Chemicals, 6th ed. Butterworth-Heineman Ltd.: Oxford 2008)and were immediately distilled before being used. Reaction developmentwas assessed by thin layer chromatography, using silica gel plates(Merck 60F254), which were visualized by UV light and developed by meansof a dissolution containing 3 g of potassium permanganate (KMnO₄), 20 gof potassium carbonate (K₂CO₃), 5 mL of 5% sodium hydroxide dissolution(NaOH 5%) and 300 mL of water (H₂O). The products were purified bypressure column chromatography on silica gel, Merck (230-400 mesh).

Example 1 Preparation of1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethylN,N-diethyldithiocarbamate (Id₂)

A solution of5-(tert-butyldiphenylsyliloxymethyl)-5-hydroxy-4-methyl-5H-furan-2-oneVIId (212 mg, 0.554 mmol) in absolute EtOH (4 mL), was added at roomtemperature (RT), a CH₃NHNH₂ solution (0.06 mL, 1.108 mmol) in absoluteEtOH (1 mL). The reaction mixture was stirred at reflux for 18 hours.Once the reaction was finished, and once the resulting solution wascooled, the solvent was removed under vacuum and the residue obtainedwas purified by column chromatography on silica gel, using hexane/ethylacetate (3:1) as eluent, thus obtaining6-(tert-butyldiphenylsyliloxymethyl)-2,5-dimethyl-3(2H)-pyridazinoneVIIId_(I) (313 mg, 52%). EMAR (ESI): m/z calculated for C₂₃H₂₉N₂O₂Si[M+H]⁺, 393.19983; found 393.19928.

¹H NMR (CDCl₃) δ: 7.68 (m, 4H,), 7.67 (m, 6H,), 6.70 (d, 1H, J=1.1 Hz),4.64 (s, 2H), 3.62 (s, 3H,), 2.34 (d, 3H, J=1.1 Hz), 1.01 (s, 9H).

A solution of the compound VIIId_(I) (74 mg, 0.188 mmol) in THF (4 mL),was added at RT, and under argon (Ar) atmosphere, a 1M TBAF solution inTHF (0.2 mL, 0.188 mmol). The reaction mixture was kept under stirring,at RT and under Ar atmosphere for 15 minutes. Once the reaction wasfinished, it was added some drops of saturated NaHCO₃ solution, stirringwas kept for 15 additional minutes and then it was dried with anhydrousNa₂SO₄. The resulting suspension was filtered and the filtrate wasconcentrated to dryness under vacuum. The residue obtained was purifiedby column chromatography on silica gel, using ethyl acetate/methanol(9.5:0.5) as eluent, obtaining6-hydroxymethyl-2,5-dimethyl-3(2H)-pyridazinone VId_(I) (26 mg, 86%).EMAR (EI): m/z calculated for C₇H₁₀N₂O₂[M]⁺, 154.0742, found 154.0735.

¹H NMR (CDCl₃) δ: 6.70 (d, 1H, J=1.1 Hz), 4.58 (s, 2H), 3.73 (s, 3H),2.21 (d, 3H, J=1.1 Hz).

A solution of the compound VId_(I) (48 mg, 0.311 mmol) in CH₂Cl₂ (8 mL)was successively added CBr₄ (207 mg, 0.623 mmol) and PPh₃ (163 mg 0.623mmol). The reaction mixture was kept under stirring at reflux, under Aratmosphere for 6 hours. Once the reaction was finished, and once theresulting solution was cooled, it was treated with a saturated NaHCO₃solution (2 mL), extracted with CH₂Cl₂ (3×5 mL) and the organic extractwas dried over anhydrous Na₂SO₄. The resulting suspension was filteredand the filtrate was vacuum concentrated. The residue obtained waspurified by column chromatography on silica gel, using ethylacetate/methanol (8.5:1.5) as eluent, thus obtaining6-bromomethyl-2,5-dimethyl-3(2H)-pyridazinone IVd_(I) (65 mg, 96%). EMAR(ESI): m/z calculated for C₇H₁₀BrN₂O, 216.99710 [M+H]⁺, found 216.99627.

¹H NMR (CDCl₃) δ:6.73 (m, 1H), 4.37 (s, 2H), 3.74 (s, 3H), 2.32 (d, 3H,J=1.1 Hz).

A diethylamine solution (8 μL, 0.077 mmol) in DMF (1 mL), was added CS₂(9 μL, 0.141 mmol) and K₃PO₄ (16 mg, 0.077 mmol). The mixture obtainedwas stirred at RT and under Ar atmosphere for 30 minutes. Then, asolution of the compound IVd_(I) (11 mg, 0.051 mmol) in DMF (1 mL) wasadded and stirring was kept under the same conditions for 22 hours.Then, the reaction mixture was treated with H₂O (0.5 mL) and wasconcentrated to dryness under vacuum. The residue obtained was purifiedby column chromatography on silica gel (eluent: hexane/ethyl acetate 3:1and 1:3) thus obtaining the compound Id₂ (14 mg, 97%). EMAR (ESI): m/z[M+H]⁺ calculated for C₁₂H₂₀N₃OS₂, 286.10423, found 286.10546.

¹H NMR (CDCl₃) δ: 6.69 (s, 1H), 4.52 (s, 2H), 4.03 (c, 2H, J=7.0 Hz),3.72 (s, 3H), 3.65 (c, 2H, J=7.0 Hz), 2.26 (s, 3H), 1.28 (m, 6H).

Example 2 Preparation of1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethylPyrrolidin-1-ylcarbodithioate (Id₃)

According to the procedure described for obtaining the compound Id₂, asolution of pyrrolidine (8 μL, 0.096 mmol), CS₂ (11 μL, 0.175 mmol) andK₃PO₄ (20 mg, 0.096 mmol) in DMF (1 mL) was treated with a solution ofthe compound IVd_(I) (10 mg, 0.046 mmol) in DMF (1 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 3:1 and hexane/ethyl acetate 1:3), obtaining thecompound Id₃ (13 mg, 100%). EMAR (ESI): m/z calculated for C₁₂H₁₈N₃₀S₂,284.08858 [M+H]⁺, found 284.08856.

¹H NMR (CDCl₃) δ: 6.69 (m, 1H), 4.55 (s, 2H), 3.94 (t, 2H, J=6.9 Hz),3.72 (s, 3H), 3.65 (t, 2H, J=6.9 Hz), 2.26 (d, 3H, J=1.1 Hz), 2.08 (m,2H), 1.99 (m, 2H).

Example 3 Preparation of1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethylPiperidin-1-ylcarbodithioate (Id₄)

According to the procedure described for obtaining the compound Id₂, asolution of piperidine (8 μL, 0.081 mmol), CS₂ (9 μL, 0.147 mmol) andK₃PO₄ (17 mg, 0.081 mmol) in DMF (1 mL) was treated with a solution ofthe compound IVd_(I) (15 mg, 0.069 mmol) in DMF (1 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 3:1, 1:1, 1:3 and 1:4), obtaining the compound Id₄(19 mg, 95%). EMAR (ESI): m/z calculated for C₁₃H₂₀N₃₀S₂, 298.10423[M+H]⁺, found 298.10379.

¹H NMR (CDCl₃) δ: 6.69 (m, 1H), 4.53 (s, 2H), 4.28 (m, 2H), 3.88 (m,2H), 3.72 (s, 3H), 2.26 (d, 3H, J=1.0 Hz), 1.70 (m, 6H).

Example 4 Preparation of1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethylmorpholin-4-ylcarbodithioate (Id₅)

According to the procedure described for obtaining the compound Id₂, asolution of morpholine (8 μL, 0.091 mmol), CS₂ (10 μL, 0.165 mmol) andK₃PO₄ (19 mg, 0.091 mmol) in DMF (1 mL) was treated with a solution ofthe compound IVd₁ (10 mg, 0.046 mmol) in DMF (1 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 3:1, 1:2, and 1:4), obtaining the compound Id₅ (13mg, 94%). EMAR (ESI): m/z calculated for C₁₂H₁₈N₃O₂S₂, 300.08349 [M+H]⁺,found 300.08357.

¹H NMR (CDCl₃) δ: 6.70 (m, 1H), 4.54 (s, 2H), 4.33 (m, 2H), 3.96 (m,2H), 3.77 (m, 4H), 3.72 (s, 3H), 2.27 (d, 3H, J=1.0 Hz).

Example 5 Preparation of1,4-dimethyl-6-oxo-1,6-dihydropyridazin-3-ylmethyl4-benzoylpiperazin-1-ylcarbodithioate (Id₁₁)

According to the procedure described for obtaining the compound Id₂, asolution of 1-benzoylpiperazine (13 mg, 0.068 mmol), CS₂ (8 μL, 0.132mmol) and K₃PO₄ (14 mg, 0.068 mmol) in DMF (1 mL) was treated with asolution of the compound IVd_(I) (10 mg, 0.046 mmol) in DMF (1 mL). Theresidue obtained was purified by column chromatography on silica gel(eluent: hexane/ethyl acetate 1:1, 1:2 and 1:4), obtaining the compoundId₁₁ (18 mg, 97%). EMAR (ESI): m/z calculated for C₁₉H₂₃N₄O₂S₂,403.12569 [M+H]⁺, found 403.12569.

¹H NMR (CDCl₃) δ: 7.43 (m, 5H), 6.71 (s, 1H), 4.53 (s, 2H), 4.18 (m,4H), 3.84 (m, 2H), 3.72 (s, 3H), 3.65 (m, 2H), 2.26 (s, 3H).

Example 6 Preparation of2-(1-methyl-6-oxo-1,6-dihydropyridazin-3-yl)ethylPiperidin-1-ylcarbodithioate (Ia₁₅)

According to the procedure described for IVd_(I), a solution of6-(2-hydroxyethyl)-2-methyl-3(2H)-pyridazinone VIa_(II) (6 mg, 0.039mmol) in CH₂Cl₂ (6 mL) was treated with CBr₄ (26 mg, 0.078 mmol) andPPh₃ (20 mg 0.078 mmol). The residue obtained was purified by columnchromatography on silica gel using methylene chloride/methanol as eluent(88:2), obtaining 6-(2-bromoethyl)-2-methyl-3(2H)-pyridazinone IVa_(II)(7 mg, 83%).

¹H NMR (CDCl₃) (δ: 7.14 (d, 1H, J=9.4 Hz), 6.90 (d, 1H, J=9.4 Hz), 3.76(s, 3H), 3.64 (t, 2H, J=7.0 Hz), 3.14 (t, 2H, J=7.0 Hz).

According to the procedure described for obtaining the compound Id₂, asolution of piperidine (8 μL, 0.081 mmol), CS₂ (9 μL, 0.147 mmol) andK₃PO₄ (17 mg, 0.081 mmol) in DMF (1 mL) was treated with a solution ofthe compound IVa_(II) (10 mg, 0.046 mmol) in DMF (1 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 3:1, 1:1 and 1:3), obtaining the compound Ia₁₅ (8.5mg, 89%). EMAR (ESI): m/z calculated for C₁₃H₂₀N₃OS₂, 298.10423 [M+H]⁺,found 298.10432.

¹H NMR (CDCl₃) δ: 7.25 (d, 1H, J=9.5 Hz), 6.89 (d, 1H, J=9.5 Hz), 4.29(m, 2H), 3.86 (m, 2H), 3.75 (s, 3H), 3.59 (t, 2H, J=7.4 Hz), 3.00 (t,2H, J=7.4 Hz), 1.70 (m, 6H).

Example 7 Preparation of2-(1-methyl-6-oxo-1,6-dihydropyridazin-3-yl)propylPyrrolidin-1-ylcarbodithioate (Ia₂₅)

According to the procedure described for IVd_(I), a solution of6-(2-hydroxypropyl)-2-methyl-3(2H)-pyridazinone VIa_(II) (6 mg, 0.036mmol) in CH₂Cl₂ (6 mL) was treated with CBr₄ (30 mg, 0.096 mmol) andPPh₃ (30 mg 0.114 mmol). The residue obtained was purified by columnchromatography on silica gel using methylene chloride/methanol as eluent(88:2), obtaining 6-(2-bromopropyl)-2-methyl-3(2H)-pyridazinone IVa_(II)(7 mg, 84%). EMAR (EI): m/z calculated for C₈H₁₁BrN₂O, 230.0055 [M]⁺;found 230.0057.

¹H NMR (CDCl₃) δ: 7.11 (d, 1H, J=9.6 Hz), 6.89 (d, 1H, J=9.6 Hz), 3.75(s, 3H), 3.47 (t, 2H, J=6.4 Hz), 2.76 (t, 2H, J=7.4 Hz), 2.22 (m, 2H).

According to the procedure described for obtaining the compound Id₂, asolution of pyrrolidine (8 μL, 0.096 mmol), CS₂ (9 μL, 0.147 mmol) andK₃PO₄ (17 mg, 0.081 mmol) in DMF (1 mL) was treated with a solution ofthe compound IVa_(II) (9 mg, 0.039 mmol) in DMF (1 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 1:1, 1:2), obtaining the compound Ia₂₅ (11 mg,95%). EMAR (ESI): m/z calculated for C₁₃H₂₀N₃OS₂, 298.10478 [M+H]⁺,found 298.10432.

¹H NMR (CDCl₃) δ: 7.13 (d, 1H, J=9.5 Hz), 6.89 (d, 1H, J=9.5 Hz), 3.93(t, 2H, J=6.9 Hz), 3.75 (s, 3H), 3.64 (t, 2H, J=6.8 Hz), 3.36 (t, 2H,J=7.3 Hz), 2.71 (t, 2H, J=7.6 Hz), 2.07 (m, 4H), 1.98 (m, 2H).

Example 8 Preparation of 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethylN,N-diethyldithiocarbamate (IIc₂)

A solution of 3-(tert-butyldiphenylsyliloxymethyl)furan (3.00 g, 8.92mmol) in dry MeOH (40 mL) was added diisopropylethylamine (7 mL, 40.20mmol) and rose Bengal (15 mg) and was purged at RT with O₂ for 1 h. Thereaction mixture was cooled at −78° C. and irradiated with a 200 W lampunder an O₂ atmosphere for 5 h. Then it was left to reach RT and thesolvent was removed under vacuum. The residue obtained was dissolved inCH₂Cl₂ (40 mL) and a 0.12 M oxalic acid solution (350 mL) was added,stirring for 30 min. The resulting mixture was extracted with CH₂Cl₂(3×100 mL) and the combined organic phases were dried, filtered andconcentrated under vacuum. The residue obtained was purified by columnchromatography on silica gel (eluent: ethyl acetate/hexane 1:2), with amixture of 4-(tert-butyldiphenylsyliloxymethyl)-5-hydroxy-5H-furan-2-oneXI₁ and of 3-(tert-butyldiphenylsyliloxymethyl)-5-hydroxy-5H-furan-2-oneXV₁ (3.29 g, 100%) being isolated at a ratio 4:1. EMAR (ESI): m/zcalculated for C₂₁H₂₅O₄Si, 369.15166 [M+1]; found 369.15162.

A solution of the mixture of compounds XI₁ y XV₁ (531 mg, 1.44 mmol, 4:1ratio) in absolute ethanol (15 mL), was added benzylhydrazinedihydrochloride (BnNHNH₂.2HCl, 562 mg, 2.88 mmol) and Et₃N (0.6 mL, 4.30mmol). The reaction mixture was stirred at reflux for 7 h. Then, thesolvent was removed under vacuum, and the residue obtained was purifiedby column chromatography on silica gel (eluent: hexane/ethyl acetate9:1), isolating2-benzyl-4-(tert-butyldiphenylsyliloxymethyl)pyridazin-3(2H)-one XVIc₁(43 mg, 32%) and then2-benzyl-5-(tert-butyldiphenylsyliloxymethyl)pyridazin-3(2H)-one XIIc₁(277 mg, 53%).

2-Benzyl-4-(tert-butyldiphenylsyliloxymethyl)pyridazin-3(2H)-one XVIc₁EMAR (ESI): m/z calculated for C₂₈H₃₁N₂O₂Si, 455.21493 [M+1]; found455.21371.

¹H-NMR (CDCl₃, δ): 7.85 (d, 1H, J=4.0 Hz), 7.63 (m, 4H), 7.51 (m, 1H),7.36 (m, 11H), 5.29 (s, 2H), 4.73 (d, 2H, J=1.5 Hz), 1.12 (s, 9H).

2-Benzyl-5-(tert-butyldiphenylsyliloxymethyl)pyridazin-3(2H)-one XIIc₁EMAR (ESI): m/z calculated for C₂₈H₃₁N₂O₂Si, 455.21493 [M+1]; found455.21473.

¹H-NMR (CDCl₃, δ): 7.64 (m, 5H, H6), 7.37 (m, 11H), 6.96 (m, 1H), 5.32(s, 2H), 4.55 (d, 2H, J=1.5 Hz), 1.09 (s, 9H).

According to the procedure described for VId, a2-benzyl-5-(tert-butyldiphenylsyliloxymethyl)pyridazin-3(2H)-one XIIc₁solution (247 mg, 0.54 mmol) in THF (5 mL), was treated with a 1M TBAFsolution in THF (0.8 mL, 0.81 mmol). The residue obtained was purifiedby column chromatography on silica gel (eluent: ethyl acetate/methanol98:2), obtaining 2-benzyl-5-hydroxymethylpyridazin-3(2H)-one Xc_(I) (102mg, 87%). EMAR (ESI): m/z calculated for C₁₂H₁₃N₂O₂, 217.09715 [M+1];found 217.09782.

According to the procedure described for IVd_(I), a2-benzyl-5-hydroxymethylpyridazin-3(2H)-one Xc_(I) solution (83 mg, 0.39mmol) in CH₂Cl₂ (5 mL), was treated with CBr₄ (256 mg, 0.77 mmol) andPPh₃ (202 mg, 0.77 mmol). The residue was purified by columnchromatography on silica gel (eluent: dichloromethane/methanol99.5:0.5), obtaining 2-benzyl-5-bromomethylpyridazin-3(2H)-one IXc_(I)(62 mg, 58%). EMAR (ESI): m/z calculated for C₁₂H₁₂BrN₂O, 279.01275[M+1]; found 279.01210.

¹H-NMR (CDCl₃, δ): 7.77 (d, 1H, J=1.9 Hz), 7.40 (m, 2H), 7.29 (m, 3H),6.85 (d, 1H, J=1.9 Hz), 5.30 (s, 2H), 4.17 (s, 2H).

According to the procedure described for obtaining the compound Id₂, adiethylamine solution (3.9 μL, 0.037 mmol), CS₂ (4.1 μL, 0.068 mmol) andK₃PO₄ (8 mg, 0.037 mmol) in DMF (2 mL) was treated with a solution ofthe compound IXc_(I) (9.5 mg, 0.034 mmol) in DMF (2 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 4:1, 3:1, 2:1), obtaining the compound IIc₂ (10.2mg, 86%). EMAR (ESI): m/z [M+H]⁺ calculated for C₁₇H₂₂N₃OS₂, 348.12043,found 348.11976.

¹H NMR (CDCl₃) δ:7.84 (m, 1H), 7.43 (m, 2H), 7.32 (m, 3H), 6.91 (m, 1H),5.30 (s, 2H), 4.45 (s, 2H), 4.03 (c, 2H, J=6.5 Hz), 3.76 (c, 2H, J=6.5Hz), 1.30 (m, 6H).

Example 9 Preparation of 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethylPyrrolidin-1-ylcarbodithioate (IIc₃)

According to the procedure described for obtaining the compound Id₂, apyrrolidine solution (3.2 μL, 0.038 mmol), CS₂ (4.1 μL, 0.068 mmol) andK₃PO₄ (8 mg, 0.037 mmol) in DMF (2 mL) was treated with a solution ofthe compound IXc_(I) (10 mg, 0.035 mmol) in DMF (2 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 3:1, 2:1), obtaining the compound IIc₃ (11.7 mg,97%). EMAR (ESI): m/z calculated for C₁₇H₂₀N₃OS₂, 348.10478 [M+H]⁺;found 346.19423.

¹H NMR (CDCl₃) δ: 7.83 (d, 1H, J=2.1 Hz), 7.40 (m, 2H), 7.29 (m, 3H),6.89 (m, 1H), 5.28 (s, 2H), 4.43 (s, 2H), 3.91 (t, 2H, J=6.9 HZ), 3.65(t, 2H, J=6.9 Hz), 2.09 (q, 2H, J=6.9 Hz), 2.00 (q, 2H, J=6.9 Hz).

Example 10 Preparation of 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethylPiperidin-1-ylcarbodithioate (IIc₄)

According to the procedure described for obtaining the compound Id₂, apiperidine solution (3.1 μL, 0.031 mmol), CS₂ (3.5 μL, 0.057 mmol) andK₃PO₄ (8 mg, 0.037 mmol) in DMF (2 mL) was treated with a solution ofthe compound IXc_(I) (10 mg, 0.035 mmol) in DMF (2 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 4:1, 3:1, 2:1), obtaining the compound IIc₄ (9.4mg, 90%). EMAR (ESI): m/z calculated for C₁₈H₂₂N₃OS₂, 360.12043 [M+H]⁺;found 360.11968.

¹H NMR (CDCl₃) δ: 7.82 (d, 1H, J=2.3 Hz), 7.41 (m, 2H), 7.29 (m, 3H),6.88 (m, 1H), 5.28 (s, 2H), 4.44 (s, 2H), 4.26 (m, 2H), 3.87 (m, 2H),1.71 (m, 6H).

Example 11 Preparation of 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethylMorpholin-4-ylcarbodithioate (IIc₅)

According to the procedure described for obtaining the compound Id₂, amorpholine solution (3.4 μL, 0.039 mmol), CS₂ (4.4 μL, 0.072 mmol) andK₃PO₄ (8 mg, 0.037 mmol) in DMF (2 mL) was treated with a solution ofthe compound IXc_(I) (10 mg, 0.035 mmol) in DMF (2 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 4:1, 3:1, 2:1), obtaining the compound IIc₅ (12 mg,92%). EMAR (ESI): m/z calculated for C₁₇H₂₀N₃O₂S₂, 362.09969 [M+H]⁺;found 362.09914.

¹H NMR (CDCl₃) δ:7.80 (d, 1H, J=2 Hz), 7.41 (m, 2H), 7.30 (m, 3H), 6.89(m, 1H), 5.28 (s, 2H), 4.45 (s, 2H), 4.30 (m, 2H), 3.93 (m, 2H), 3.77(m, 4H).

Example 12 Preparation of 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethyl4-benzoylpiperazin-1-ylcarbodithioate (IIc₁₁)

According to the procedure described for obtaining the compound Id₂, abenzoylpiperazine solution (7 mg, 0.038 mmol), CS₂ (4.1 μL, 0.068 mmol)and K₃PO₄ (8 mg, 0.037 mmol) in DMF (2 mL) was treated with a solutionof the compound IXc_(I) (10 mg, 0.035 mmol) in DMF (2 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 2:1, 1:1), obtaining the compound IIc₁₁ (12 mg,74%). EMAR (ESI): m/z calculated for C₂₄H₂₅N₄O₂S₂, 465.14189 [M+H]⁺;found 465.14134.

¹H NMR (CDCl₃) δ: 7.89 (d, 1H, J=2.3 Hz), 7.43 (m, 7H), 7.30 (m, 3H),6.89 (m, 1H), 5.28 (s, 2H), 4.44 (s, 2H), 4.39-3.54 (m, 8H).

Example 13 Preparation of 1-benzyl-6-oxo-1,6-dihydropyridazin-5-ylmethylPyrrolidin-1-ylcarbodithioate (IIIc₃)

According to the procedure described for VId_(I), a2-benzyl-4-(tert-butyldiphenylsyliloxymethyl)pyridazin-3(2H)-oneXVIc_(I) solution (70 mg, 0.15 mmol) in THF (5 mL) was treated with a 1MTBAF solution in THF (0.2 mL, 0.23 mmol). The residue obtained waspurified by column chromatography on silica gel (eluent: ethylacetate/methanol 98:2), obtaining2-benzyl-4-hydroxymethylpyridazin-3(2H)-one XIVc_(I) (25 mg, 75%). EMAR(ESI): m/z calculated for C₁₂H₁₃N₂O₂, 217.09715 [M+1]; found 217.09651.

According to the procedure described for IVd_(I), a2-benzyl-4-hydroxymethylpyridazin-3(2H)-one XIVc_(I) solution (29 mg,0.13 mmol) in CH₂Cl₂ (5 mL) was treated with CBr₄ (90 mg, 0.27 mmol) andPPh₃ (70 mg, 0.27 mmol). The residue was purified by columnchromatography on silica gel (eluent: hexane/ethyl acetate 6:1)obtaining 2-benzyl-4-bromomethylpyridazin-3(2H)-one XIIIc_(I) (31 mg,84%).

¹H NMR (CDCl₃) δ: 7.77 (d, 1H, J=4 Hz), 7.46-7.41 (m, 2H), 7.36-7.27 (m,4H), 5.35 (s, 2H), 4.39 (s, 2H).

According to the procedure described for obtaining the compound Id₂, apyrrolidine solution (3.9 μL, 0.046 mmol), CS₂ (4.4 μL, 0.072 mmol) andK₃PO₄ (8 mg, 0.037 mmol) in DMF (2 mL) was treated with a solution ofthe compound XIIIc_(I) (10 mg, 0.035 mmol) in DMF (2 mL). The residueobtained was purified by column chromatography on silica gel (eluent:dichloromethane/methanol 98:2), obtaining the compound IIIc₃ (6.6 mg,55%). EMAR (ESI): m/z calculated for C₁₇H₂₀N₃OS₂, 346.10478 [M+H]⁺;found 346.10451.

¹H NMR (CDCl₃) δ: 7.70 (d, 1H, J=4.1 Hz), 7.51 (d, 1H, J=4.1 Hz)7.41-4.43 (m, 2H), 7.33-7.26 (m, 3H), 5.32 (s, 2H), 4.54 (s, 2H), 3.90(t, 2H, J=6.7 HZ), 3.63 (t, 2H, J=6.7 Hz), 2.05 (q, 2H, J=6.7 Hz), 1.95(q, 2H, J=6.7 Hz).

Example 14 Preparation of 1-benzyl-6-oxo-1,6-dihydropyridazin-5-ylmethylMorpholin-4-ylcarbodithioate (IIIc₅)

According to the procedure described for obtaining the compound Id₂, amorpholine solution (7 μL, 0.080 mmol), CS₂ (10 μL, 0.160 mmol) andK₃PO₄ (17 mg, 0.079 mmol) in DMF (2 mL) was treated with a solution ofthe compound XIIIc_(I) (11 mg, 0.040 mmol) in DMF (2 mL). The residueobtained was purified by column chromatography on silica gel (eluent:hexane/ethyl acetate 3:1), obtaining the compound IIIc₅ (14 mg, 97%).

EMAR (ESI): m/z calculated for C₁₇H₂₀N₃O₂S₂, 362.09969 [M+H]⁺; found362.09914.

¹H NMR (CDCl₃) δ: 7.71 (d, 1H, J=4.1 Hz), 7.47 (d, 1H, J=4.1 Hz), 7.42(d, 2H, J=6.8 Hz), 7.34-7.26 (m, 3H), 5.31 (s, 2H), 4.55 (s, 2H),4.38-4.23 (m, 2H), 4.11-3.98 (m, 2H), 3.81-3.73 (m, 4H).

Example 15 Preparation of 1-benzyl-6-oxo-1,6-dihydropyridazin-4-ylmethyl4-benzoylpiperazin-1-ylcarbodithioate (IIIc₁₁)

According to the procedure described for obtaining the compound Id₂, asolution of benzoylpiperazine (11.4 mg, 0.060 mmol), CS₂ (7 μL, 0.112mmol) and K₃PO₄ (13 mg, 0,060 mmol) in DMF (2 mL), was treated with asolution of the compound XIIIc_(I) (9 mg, 0.030 mmol) in DMF (2 mL). Theresidue obtained was purified by column chromatography on silica gel(eluent: dichloromethane, dichloromethane/methanol 99:1), obtaining thecompound IIIc₁₁ (14 mg, 100%).

EMAR (ESI): m/z calculated for C₂₄H₂₅N₄O₂S₂, 465.14189 [M+H]⁺; found465.14071.

¹H NMR (CDCl₃) δ: 7.73 (d, 1H, J=4.0 Hz), 7.48 (d, 1H, J=4.0 Hz), 7.42(m, 7H), 7.31 (m, 3H), 5.33 (s, 2H), 4.56 (s, 2H), 4.18 (m, 4H), 3.83(m, 2H), 3.62 (m, 2H).

Inhibition of MAOs

Determination of MAO Isoforms Activity

The effects of compounds of formulas I, II and III on monoamine oxidasewere determined by measuring the production of hydrogen peroxide (H₂O₂),and therefore the production of resorufin from p-tyramine, a substratecommon to both isoenzymes (MAO-A and MAO-B). This was performed by usingAmplex® Red reagent (Molecular Probes, Eugene, Oreg., USA) and MAOisoforms present in the microsomal fraction, prepared from insect cells(BTI-TN-5B1-4) infected with recombinant baculovirus, containing humanMAO-A or MAO-B cDNA (Sigma-Aldrich Química S.A., Alcobendas, Spain).

Production of H₂O₂ catalysed by the 2 MAO isoforms can be detected byusing Amplex® Red reagent (10-acetyl-3,7-dihydroxyphenoxazine), anon-fluorescent substance, highly sensitive, which reacts with H₂O₂ inthe presence of horseradish peroxidase for producing a fluorescentproduct, resorufin.

In our experiments, the MAO activity was assessed with the abovementioned method, adapting the general process previously described(Biochem. Biophys. Res. Comm. 344, 688-695, 2006).

In the first place, 0.1 ml of sodium phosphate buffer (0.05 M, pH 7.4)was incubated, containing different concentrations of the novelcompounds under study (or reference inhibitors) and the amount of humanrecombinant MAO-A or MAOB required for obtaining in our experimentalconditions the same reaction speed in the presence of both isoenzymes;that is, for oxidizing, in the absence of drugs (control group), 165pmoles of p-tyramine per minute (MAO-A: 1.1 μg; specific activity: 150nmoles of p-tyramine oxidized to p-hydroxyphenylacetaldehyde per minuteper protein mg; MAO-B: 7.5 μg; specific activity: 22 nmoles ofp-tyramine transformed per minute per protein mg). Said incubation wasperformed for 15 minutes at 37° C. in 96 well plates with black and flatbottom (Microtest™ plate, BD, Franklin Lakes, N.J., USA), already placedin the light-tight chamber of the fluorescence reader (see the modelbelow). After incubation period, reaction was started by adding (finalconcentrations) 200 μM of Amplex® Red reagent, 1 unit (U)/ml ofhorseradish peroxidase and 1 mM of p-tyramine as substrate, both forstudies carried out with MAO-A and those carried out with MAO-B.

H₂O₂ and, accordingly, resorufin production was quantified at 37° C. ina plate fluorescence reader (FLX800™, Bio-Tek® Instruments, Inc.,Winooski, Vt., EE.UU.), determining the fluorescence generated(excitation 545 nm, emission 590 nm) for 15 minutes, a period in whichthe increase of fluorescence was linear from the beginning.

Simultaneously, control experiments were conducted substituting thedrugs (compounds of formulas I, II and III or reference inhibitors) withthe appropriate vehicle dilutions. Furthermore, it was determined thepossible capacity of the drugs for modifying the fluorescence generatedin the reaction mixture by a non-enzymatic inhibition (for example, bydirect reaction with the Amplex® Red reagent), and for that reason thedrugs were added to solutions containing only the Amplex® Red reagent ina sodium phosphate buffer.

Specific fluorescence emission was calculated (used for obtaining thefinal results) after subtracting the background activity, beingdetermined in vials in which solutions with MAO isoforms weresubstituted by sodium phosphate buffer.

Statistical Data and Analysis Presentation

Unless otherwise indicated, the results shown in the text and in thetables are expressed as the mean±mean standard error (m.s.e.) of fiveexperiments. The statistically significant difference between two means(P<0.05 or P<0.01) was determined by one-way analysis of variance(ANOVA), followed by Dunnett's multiple comparison test.

In order to study the possible effects of the compounds of formulas I,II and III, and of the reference inhibitors, about the enzymaticactivity of the MAO isoforms, fluorescence per time unit was assessed(quantified as random fluorescence/minute units) and indirectly H₂O₂production; and accordingly, the pmoles/min of resorufin produced in thereaction between H₂O₂ and Amplex® Red reagent. In order to achieve that,several concentrations of resorufin were used with the purpose ofcreating a standard curve, being X=pmoles of resorufin andY=fluorescence random units. The pmoles of resorufin produced areequivalent to the pmoles of oxidised p-tyramine, since stoichiometry ofthe reaction is 1:1.

In these experiments, MAOI activity of the compounds of formula I, IIand III and that of the reference inhibitors is expressed as IC₅₀, thatis, the required concentration of each compound for producing areduction in the control value of the MAO isoforms enzymatic activity by50%. For determining the IC₅₀ of each compound the computer programOrigin™ 5.0 (Microcal Software, Inc., Northampton, Mass., USA) was used.The IC₅₀ values were calculated from the straight lines equationsobtained by linear regression (method of the least squares) of thepoints resulting from representing the log of the molar concentration ofthe compound studied (axis of abscissas) against the percentage of thecontrol MAO activity inhibition achieved with said concentration (axisof ordinates). This linear regression was performed using for eachcompound the data obtained with 4 to 6 concentrations capable ofinhibiting between 20% and 80% of the control enzymatic activity of MAOisoenzymes. Furthermore, the ratio was calculated [IC₅₀ (MAO-A)]/[IC₅₀(MAO-B)] as an indicator of selectivity in the inhibition shown on bothisoforms.

Drugs and Chemical Compounds

The drugs and chemical substances used in the experiments were compoundsof formulas I, II and III, moclobemide (kindly provided by Hoffman-LaRoche Laboratories, Basel, Switzerland), selegiline and iproniazidphosphate (acquired in Sigma-Aldrich, Spain), resorufin sodium salt,clorgiline hydrochloride, p-tyramine hydrochloride, sodium phosphate andhorseradish peroxidase (provided in the MAO assay kit Amplex® Red ofMolecular Probes).

Appropriate dilutions of the above compounds were prepared in Milli-Q®water (Millipore Ibérica S.A., Madrid, Spain) every day before usingthereof, from the following concentrated stock solutions kept at −20°C.: compounds of formulas I, II and III (0.1 M) in dimethyl sulphoxide(DMSO, Sigma-Aldrich); selegiline, moclobemide, iproniazid, resorufin,clorgiline, p-tyramine and horseradish peroxidase (0.1 M) in Milli-Q®water.

Due to photosensitivity of some of the substances being used (forexample, Amplex® Red reagent), all experiments were performed indarkness. In any of the assays, neither Milli-Q® water nor the vehiclebeing used (DMSO) showed a significant pharmacological effect.

Results

The compounds used for the present invention of the general formula I,II and III are selective inhibitors of MAO-B. Table XI show the IC₅₀values in micromoles/L (μM) of the compounds detailed above (Id₂, Id₃,Id₄, Id₅, Id₁₁, Ia₁₅, Ia₂₅, IIc₂, IIc₃, IIc₄, IIc₅, IIc₁₁, IIIc₃, IIIc₅,IIIc₁₁).

TABLE XI IC₅₀ values of the studied compounds (including referenceinhibitors) on enzymatic activity of human recombinant MAO isoforms andselectivity index for MAO-B ([IC₅₀ (MAO-A)]/[IC₅₀ (MAO-B)]). CompoundIC₅₀ hMAO-A (μM) IC₅₀ hMAO-B (μM) S.I. Id₂ *** *** — Id₃ ** * — Id₄ *** 7.48 ± 0.34 13.4  Id₅ *** 38.57 ± 1.74  2.6 Id₁₁ *** 44.53 ± 2.00  2.2Ia₁₅ ** 11.88 ± 0.53 >8.4^(b) Ia₂₅ ** 66.49 ± 4.43 >1.5^(b) IIc₂ **44.25 ± 2.95 >2.3^(b) IIc₃ ***  9.68 ± 0.65 10.3  IIc₄ **  6.71 ± 0.45>15^(b )  IIc₅ ** *** — IIc₁₁ ** ** — IIIc₃ ** 33.96 ± 2.26 >2.9^(b)IIIc₅ ** 24.05 ± 1.60 >4.2^(b) IIIc₁₁ ** ** — Clorgiline 0.0052 ±0.00092^(a) 63.41 ± 1.20     0.000082 Selegiline 68.73 ± 4.21^(a)  0.017 ± 0.0019   4.043 Iproniazid 6.56 ± 0.76   7.54 ± 0.36   0.87Moclobemide 361.38 ± 19.37    *  <0.36^(b) Each IC₅₀ value is the mean ±mean standard deviation of 5 experiments (n = 5). ^(a)P < 0.01 withrespect to the corresponding IC₅₀ value obtained against MAO-B,determined by ANOVA/Dunnett's test. ^(b)Value calculated considering asIC50 against MAO-A or MAO-B the highest concentration studied (100 μM or1 mM). * Inactive at 1 mM (highest concentration studied) ** Inactive at100 μM (highest concentration assayed). At higher concentrations thecompound precipitates. *** At 100 μM inhibit enzymatic activity by45-50%. At higher concentrations the compound precipitates. SI:Selectivity index hMAO-B = IC₅₀ (hMAO-A)/IC₅₀ (hMAO-B)

Most of the compounds of the general formula I, II and III detailed inthe table are inactive against MAO-A and inhibit MAO-B with IC₅₀ valuesin the micromolar range.

IC₅₀ values of the compounds of the general formula I, II and IIIagainst MAO-B are comparable to those exhibited by some of the referenceinhibitors used in the study, such as for example iproniazid(MAO-A/MAO-B dual inhibitor), but having higher MAO-B selectivityindexes.

The results obtained indicate that MAO-B activity and selectivity of thecompounds of general formula I, II and III is more influenced by thetype of amine present in the dithiocarbamate moiety than by the positionand magnitude of the alkyl chain, and by the nature of the substituentin N of the pyridazinone ring.

Therefore, incorporating dithiocarbamate moieties to the pyridazinonering into position 4, 5 or 6 through an alkyl chain of variable length,which results in the compounds of general formula I, II and III,provides selective MAO-B inhibitors the structure of which is highlynovel for this type of activity, since no pyridazinone derivatives areknown which act as selective inhibitors of MAO isoform B.

The invention claimed is:
 1. A compound of formula (I), (II) or (III):

wherein, n is an integer number selected from 1, 2, 3, 4, 5, 6, 7, 8; Ris selected from H, —C₁-C₆ alkyl, —C₁-C₆ carboxyalkyl, —C₁-C₆ haloalkyl,—C₆-C₁₂ aryl, —C₆-C₁₂ aralkyl or —C₄-C₁₂ heteroaryl; R¹ is selected fromH, —C₁-C₆ alkyl or an halogen; R² is selected from H, —C₁-C₆ alkyl or anhalogen; R³, R⁴ are independently selected from H, —C₁-C₆ alkyl,saturated —C₁-C₆ heterocycloalkyl, —C₆-C₁₂ aryl, —C₆-C₁₂ aralkyl or—C₄-C₁₂ heteroaryl; or R³ and R⁴ form a cycle selected from C₅-C₈cycloalkyl, C₅-C₈ heterocycloalkyl, N-alkyl substituted C₅-C₈heterocycloalkyl, N-aryl substituted C₅-C₈ heterocycloalkyl,N-cycloalkyl substituted C₅-C₈ heterocycloalkyl, N-aralkyl substitutedC₅-C₈ heterocycloalkyl or N-acyl substituted C₅-C₈ heterocycloalkyl; andpharmaceutically acceptable salts thereof.
 2. The compound according toclaim 1, wherein n is an integer number selected from 1, 2, or
 3. 3. Thecompound according to claim 1, wherein R is a group selected frommethyl, phenyl or benzyl.
 4. The compound according to claim 1, whereinR¹ is a hydrogen atom.
 5. The compound according to claim 1, where R² isa hydrogen atom or a methyl group.
 6. The compound according to claim 1,wherein R³ and R⁴ form a group selected from the group consisting of:


7. The compound according to claim 1, selected from the group consistingof:


8. The pharmaceutical composition comprising a compound of formula (I),(II) or (III) according to claim 1, a pharmaceutically acceptablecarrier and at least one pharmaceutically acceptable excipient.
 9. Thepharmaceutical composition according to claim 8, comprising at least oneadditional therapeutic agent.
 10. A method of treating disorders derivedfrom MAO-B hyperactivity comprising administering a compound of formula(I), (II), or (III) according to claim 1 to a mammal in need thereof,wherein the disorder is selected from Parkinson, Alzheimer, seniledementia or ataxia.
 11. A method for the synthesis of a compound offormula (I), (II) or (III) according to claim 1, comprising a stage asdescribed in scheme 1, wherein 6(5) (4)-bromoalkyl-3(2H)-pyridazinone offormula (IV), (IX) or (XIII), a secondary amine of formula V and carbondisulphide (CS₂) react in the presence of a base in a solvent at roomtemperature


12. The method according to claim 11, wherein the solvent isdimethylformamide (DMF) and the base is K₃PO₄.